This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A fundamental gap exists in the understanding of how differences in innate host defense peptide (HDP) poten-cy and repertoires are linked to the remarkable variability in infectious disease susceptibility observed among species. Until filled, this gap constitutes a major obstacle because an appreciation of the genetic profile that renders certain species with highly potent HDPs and superior resistance towards pathogens will remain unclear. The long-term goal is to understand how the molecular profiles of scavenger HDPs can be applied towards development of novel immunotherapeutics and potent, broad-spectrum natural antibiotics with low resistance. The objective of the application is to pursue this goal by conducting comparative analysis of the salivary gland transcriptome and peptidome in wild versus captive spotted hyenas (Crocuta crocuta) exposed to different immunomodulators. The hyena is an excellent model for this study, because it exhibits unique wound-healing capabilities, and survives successfully on a diet containing extreme microbial loads, including otherwise lethal pathogens. The central hypothesis is that the immunological robustness in the spotted hyena is explained by broad-spectrum antimicrobial and immunomodulatory activity of epithelial-derived HDPs. This hypothesis has been formulated based upon preliminary data obtained by the applicant. The rationale is twofold: HDPs responsible for hyena saliva antimicrobial activity will first be identified via comparative peptidome-analysis of wild vs. captive animals, and transcriptome analysis will then illustrate differential gene-expression after immunostimulation. Manipulation of HDP signaling-pathways and/or use of gene products will thereafter be applied towards development of innovative therapies for infectious/inflammatory disorders. Directed by the preliminary data, the hypothesis will be tested via pursuit of two specific aims: 1) Isolation and characterization of low-molecular weight salivary gland peptides that exhibit distinct antimicrobial activity;and 2) Identification of immune-induced genes expressed by acinar cells after local and systemic administration of an innate immune response elicitor. Under the first specific aim, the salivary peptidome will be determined by biofractionation and MALDI-TOF mass spectroscopic analysis. Under the second specific aim, and after completion of de novo assembly of the transcriptome, acinar salivary gland cells will be isolated by fluorescence-activated cell sorting and HDPs genes analyzed under control and experimental conditions using customized SuperArrays. The methods are all feasible in the hands of the applicant or the collaborators. The approach is innovative, because it utilizes a species with very low natural susceptibility to infectious diseases to identify which HDPs are linked to immunological superiority. The proposed research is significant, because it is anticipated to markedly advance the understanding of how variation in HDP expression patterns relates to differences in infection susceptibility. Ultimately, the study can lead to development of novel biotherapeutics and low-resistance antimicrobials, and thus play a major role in treating otherwise debilitating disorders. PROJECT NARRATIVE The proposed research is of relevance to public health because knowledge on innate defense in species with low disease susceptibility may translate into development of novel biotherapeutics. The study pursuits fundamental knowledge that can be applied towards improving health, and is thus relevant to NIH's mission.